Protein Expression Prokaryotic cells S. acidocaldarius

Get tips on using pCT-NT-F2A to perform Protein Expression Eukaryotic cells - S. cerevisiae Sso7d

Get tips on using p83Xi to perform Protein Expression Eukaryotic cells - S. cerevisiae Integral membrane proteins (IMPs)

Get tips on using pFastBac-N to perform Protein Expression Eukaryotic cells - S. frugiperda AvCoV-IBV

Get tips on using pFlpBtM-II-eGFP to perform Protein Expression Eukaryotic cells - S. frugiperda EGFP

Get tips on using BacPAK6-ΔChi/Cath to perform Protein Expression Eukaryotic cells - S. frugiperda β-gal

Get tips on using pYES2-αAI-OPT to perform Protein Expression Eukaryotic cells - S. cerevisiae αAI-OPT

Plasmid isolation is an important technique in molecular biology or any kind of genetic editing. It involves amplifying plasmids overnight by transforming them into competent bacterial cells. The desired colonies of these bacteria can then be grown in shaker cultures, at appropriate shaking speed, oxygen availability and temperature. These liquid cultures can then be ultracentrifuged to pellet the bacteria, which are then used for plasmid isolation. The bacteria are first resuspended in a buffer, then lysed, neutralized, purified in a column, eluted, precipitated with ethanol and then resuspended. During plasmid isolation, it is important to lyse cells quickly because lysing bacteria for too long may lead to irreversible denaturing of the plasmid. Usually, alkaline lysis is used for isolation because it is a mild treatment. It isolates plasmid DNA and other cell components such as proteins by breaking cells apart with an alkaline solution. Precipitation removes the proteins, and the plasmid DNA recovers with alcohol precipitation. Resuspension and lysis buffers should be mixed thoroughly in order to prevent the DNA from breaking into smaller fragments. This is because broken gDNA can reanneal and remain in the solution, without binding to the column.

Get tips on using pFastBac-MRP4-his6 to perform Protein Expression Eukaryotic cells - S. frugiperda human MRP4-his6

Get tips on using pVL1392-RIP1 8-322-His to perform Protein Expression Eukaryotic cells - S. frugiperda RIP1